Status of disease prevalence, drugs and antibiotics usage in pond-based aquaculture at Narsingdi district, Bangladesh: A major public health concern and strategic appraisal for mitigation

This research aimed to investigate the present status of disease prevalence and usage of aqua drugs for various aquaculture operations in the Narsingdi region of Bangladesh. Data were collected through the market survey, preset questionnaire interview, personal contact, and participatory rural appraisal tools. Amongst the respondents, the maximum percentages were found practicing mixed cultures of carp, tilapia, and pangas. The respondents suggested that epizootic ulcerative syndrome, saprolegniasis, streptococcosis, tail and fin rot and bacillary necrosis are common fish diseases in the area. About 140 drugs of different companies used in aquaculture for different purposes such as disease treatment, growth enhancement, water quality improvement, toxic gas removal, improvement of feed conversion ratio. Zeolite, rotenone, disinfectant, oxygen precursors, ammonia reducers, and probiotics were applied for pond preparation, water, and soil quality maintenance, while 30 different antibiotics were used for the purpose of treatment. Among the available antibiotics, oxytetracycline, ciprofloxacin, enrofloxacin, erythromycin, sulphadiazine, and trimethoprim were found extensively used by the fish farmers. Four enzymes and eighteen growth promoters were identified as being utilized to enhance digestion and boost up the production. This study elicited various issues connected with application and administration of such aqua chemicals, including farmers’ ignorance about their usage, proper doses, application methods, withdrawal period, and the human health concerns associated with their irresponsible use. However, the consequences of these chemical products to the environment, animal health, and human health required further study for the betterment of mankind.


Introduction
Aquaculture already accounts for 62.5% of the world's fish production for human consumption (FAO, 2018). Bangladesh is now one of the world's major aquaculture producers, with an annual production of 2.58 million metric tons in 2019-20, and standing as the fifth leading inland aquaculture producer globally (DoF, 2020). This sector (both culture and capture fishery) of Bangladesh produces 4.5 million metric tons of fish, contributing 3.52 % and 26.37% of the national and agricultural GDP (Gross Domestic Product), respectively. More than 12 % of people of Bangladesh are involved in fisheries and aquaculture activities on a fulland part-time basis for their subsistence (DoF, 2019). Bangladesh's aquaculture sector is expanding since the state's inland output is only marginally lower than China's. Indeed, the industry serves as a second source of export revenue for the government (Shamsuzzaman et al., 2017(Shamsuzzaman et al., , 2020. The Aquaculture industry has expanded, varied, increased, and mechanically commendable in Bangladesh over the last decades. Chemicals are, in fact, the critical components of thriving farming and have been utilized in numerous structures over centuries (Faruk et al., 2008). Aqua drugs are crucial for aquatic animal health management, pond installation, water and soil quality management, feed formulation, enrichment of natural production, reproductive manipulation, live fish transportation, growth stimulation, processing, and end-product value addition (Subasinghe et al., 1996;GESAMP, 1997).
Apart from the massive use of antibiotics, aquaculture uses a variety of medicines to keep fish healthy and produce more. Potassium permanganate, sodium chloride, malachite green, formalin, glutaraldehyde, methylene blue, and hydrogen peroxide are the most frequently utilized compounds (Plumb, 1992;Sumon et al., 2020). Sodium chloride is a traditional remedy especially for treating fungal and parasitic infections in fish (Phillips, 1996). Formalin is a versatile chemical that is used to treat fungal infections as well as flush fish and fish eggs in hatcheries. Potassium permanganate (KMnO4) is the powerful oxidizer that's been certified for use in ponds for external bacterial and protozoan infestations on skin, gills, and fins (Floyd, 1993;Plumb, 1992). Antibiotics were used in aquaculture for more than 50 years to treat bacterial infections in fish (Shamsuzzaman and Biswas, 2012). Recent research has shown that antibiotic use in aquaculture and the aquatic ecosystem has been linked to the development of antimicrobial resistance (Rahman et al., 2009). Pesticides like organophosphates, rotenone, and saponin are also employed to treat disease in aquaculture. Trichlorfon, melalhion, and diptarex are the most often used organophosphates in finfish aquaculture to prevent ectoparasitic crustacean infestation. Organophosphates' substantial neurotoxicity can have substantial negative impacts on the health safety of fish farm laborers (Alderman et al., 1994).
In Bangladesh, farmed aquatic animals were found to be infected with a variety of diseases (Karim and Stellwagen, 1998;Faruk et al., 2004;BFRI, 1999). Due to a lack of vaccination and good health management practices, disease problems in Bangladesh's aquaculture business remain unresolved. Experts advised that the fish vaccinations for disease resistance prevent death and economic loss (Assefa and Abunna, 2018), but in this region, this vaccination initiative needs a massive process, infrastructure,  Figure 2. Categories of fish cultured in the study area. Md.A. Kawsar et al. Heliyon 8 (2022) e09060 funding, and timeline (Asif et al., 2021). To combat fish diseases, farmers apply a variety of chemicals and medicines. Regrettably, monitoring drugs and antibiotics used in the country's aquaculture industry has received only a little attention, resulting in the aquaculture sector suffering. Farmers have been urged by chemists and representatives from various pharmaceutical companies to use their medicines on this occasion. However, the majority of the farmers indiscriminately apply such chemicals without understanding their requirements, efficacies, and method of administration and considering this issue; the current study was carried out to assess the status of disease prevalence and to identify the different types of chemicals and antibiotics used in the health management of fish and their purposes, dosages with potential concerns.

Materials and methods
The survey was undertaken randomly in 3 hatcheries, 7 nurseries, 90 grow-out farms, 20 medicine outlets, and 10 aquaculture specialists from various fish feed and pharmaceutical industries from January to December 2020 in the Narsingdi district of Bangladesh ( Figure 1). The study area was selected because the Narsingdi district is well-known for its semi-intensive commercial fish farming, and no study on this issue has been conducted in this area. Properly structured questionnaires were designed and pre-tested by several farmers in the surrounding areas before collecting the primary data. The questionnaire focused mainly on the status of fish farming systems, diseases prevalence, affected species and therapeutants they used. Fish diseases were identified based on physical appearance and farmer's observation about infected fish. Through photographs of clinical signs, the interviewer helped them in identifying the diseases. During pre-testing, all essential information regarding the objective's completion was given much thought. Primary data were collected by a survey of farmer households in selected sites after the final questionnaire was improved, using participatory rural appraisal techniques like direct farmer interviews, personal contact, focus group discussions, and crosschecks. Several focus group discussions were held in 6 Upazilas (sub-districts) within the Narsingdi region, with each group having 15 and 25 members. The district fisheries office, upazila fisheries office, private aquaculture professionals, and medicine shops owner provided valuable secondary data as well as the available previous literatures were reviewed extensivelywhich includes    online scientific articles, government surveys, books, international studies, and media reports on aqua drugs and antibiotics used in landbased aquaculture.
The study subsequently compiled, accumulated, and analyzed all the data in MS Excel version 2010 and represented in tabular and descriptive statistical techniques.

Ethical statement
All procedures performed in studies involving animals (fish) were in accordance with the ethical standards of the "Sylhet Agricultural University Ethical Committee". Informed consent was obtained from all individual participants included in the study.

Health management tools and culture strategies
The studied farms in this experiment are assigned into six categories. Such as, polyculture of carp, tilapia and pangas, polyculture of tilapia and pangas, polyculture of koi and shingi, mono culture of shingi, koi and all others were categorized as others ( Figure 2). In carps, monosex tilapia, and pangas polyculture, the maximum percentages were found at Belabo (~70%), Shibpur (~60%) and Narsingdi (~60%). The significant number of Vietnamese koi monoculture was recorded in Palash, Monohardi, and Raipura with a percentage of about 50, 45 and 35 respectively. Stocking density and culture duration were found varied with culture strategies. Almost all culture system had a duration of 3-6 months and used commercially manufactured feed. It was found that the stocking density of carp, tilapia and pangas were 60,540 fry/ha, monoculture of koi were 748803 fry/ha, monoculture of shingi with 741315 fry/ha, mixed culture of koi and shingi with 864867 fry/ha, and mixed culture of tilapia and pangas had 56834 fry/ha.

Major aqua drugs used in pond preparation
For pond preparation and improvement of the water quality of culture ponds, a variety of conventional and emerging chemical compounds are available on the market. For pond preparation, chemicals like zeolite, lime, biofertilizer and rotenone were applied by the farmers. Table 1 provides a list of the dose suggested by manufacturer and supplier information, of these compounds, along with their active ingredients. Lime and zeolite were found to be the most extensively (51% farmers) ( Figure 3) used chemicals. The second highest use of chemicals covered by the application of lime only by 31% of the farmers.

Chemicals used as disinfectants
In surveyed area, land-based aquaculture operations reported to experience extensive use of disinfectants. According to the company's information leaflet, Timsen, Aquakleen, Sansure, Pathonil, and many other drugs are effective in both preventing and destroying bacterial and fungal diseases, as well as destroying pathogens. Salt, and commercial disinfectant specially BKC (Benzal Konium Chloride) were found to be the most widely used (41%) disinfectant ( Figure 4) to treat bacterial and fungal infections. The disinfectants indicated in Table 2 were found in the market. Moreover, some other disinfectants were found in use but with only a small percentage, such as, commercial disinfectant (28%), salt (12%) and potassium permanganate (10%) etc.

Chemical used for oxygen supply
The study found several products with identical active ingredients but under different namesin the study area for enhancing oxygen levels in the aquaculture pond. Oxidizing agents, sodium carbonates, and hydrogen peroxide are the main bioactive constituents of those chemicals (Table 3). In the study area, farmers used 12 different oxygen enhancers such as Oxy-Ren (24%), Oxy more (18%), Oxy pond (16%), Oxy gold (16%) and many other brands in their ponds ( Figure 5).

Antibiotics for disease treatment
There were 30 antibiotics found in the study area with various trade names and utilized by fish farmers during the current investigation (Table 4). The active components of these antibiotics are mostly oxytetracycline, chlortetracycline, sulphadiazine trimethoprim, amoxicillin, and sulphamethoxazole. Oxytetracycline (26%), Erythromycin (19%), and Sulphadyazine (17%) were found to be the most commonly used antibiotics, followed by Ciprofloxacin (14%), Enrofloxacin (9%), Chlortetracycline (6%), Amoxicillin (5%), and some other antibiotics with a lower frequency of use ( Figure 6). Most farmers are found ignorant about the mode of action of a particular chemical in the current investigation. As a result, while treating a disease, they first test one chemical, and if it doesn't work, they try another. They calculate the dosages of a given chemical based on their own experiences, the instructions on the packet, if any, and the advice of chemical vendors.

Chemicals used as growth promoters
A variety of chemicals were found available in the market used in aquaculture as growth promoters and production boosters. Most of the growth promoters found in the market contain vitamins, minerals, amino acids, fatty acids, antioxidants, and prebiotics. Similarly, some others were found using for enhancing growth, spawning performance, disease resistance, and bio-availability of the feed supplements (Table 5). Current research revealed that 18 growth promoters were available in the study  area. Such as, Megavit Aqua (17%), Spa (15%), Charger Gel and Nutrigel (13%) were the most common brands of growth promoters used by the farmers (Figure 7).

Probiotics used in aquaculture
The current study showed that 42% of fish farmers applied 11 different brands of probiotics, such as Pond care (27%), Aqua Star Pond (19%), Profs (17%), and Safegut (14%) and many others ( Figure 8) to control diseasecausing bacteria, adsorb toxic gases, improve water and soil quality parameters, and promote the proliferation of beneficial microbes. According to respective probiotics companies, they include a variety of beneficial bacteria at different concentrations, such as Bacillus sp., Rhodococcus sp., Rodobacter sp., Streptococcus faecalis, and many others (Table 6).

Chemicals used for obnoxious gas removal
A variety of toxic gas elimination agents are used by the farmers in their culture ponds. In this study 42% of the farmers used Yucca plant extract to remove toxic gas, 33% used a combination of Yucca and Bacillus sp., 21% just Bacillus sp., and 4% used other chemicals (Figure 9). About sixteen toxic gas removers with various trade names were identified (Table 7).

Antiparasitic agents used in aquaculture
The farmers applied eight brands of antiparasitic agents such as Verkil vet, Acimec1% solution, Delitrix etc. in their pond to kill various types of harmful external parasites in the research area (Table 9). The primary active ingredients of the available antiparasitics are ivermectin, deltamethrin, cypermethrin, and trichlorfon which are used by 47%, 37%, 7%, and 6% farmers, respectively ( Figure 11).

Common diseases reported
The current investigation identified major clinical signs in diseased fish were red spots and lesions on the body, pop-eyes, abdominal distension, ulcers on the tail, fin, and lower abdominal side. Pop-eyes, reddish ulcers, cotton like fungal growth and abdomen distension were found commonly in Carp, Tilapia, Pangas, koi and shingi. In the study area, the most prevalent diseases were epizootic ulcerative syndrome (90%), saprolegniasis (90%), streptococcosis (80%), fin and tail rot (70%), and bacillary necrosis (60%) with some other diseases being observed with lower incidence (Table 10).

Discussion
Aquaculture is becoming more commercialized and intensive day by day in Bangladesh (Shamsuzzaman et al., 2017). Various kinds of drugs are becoming an essential element of effective aquaculture production. The goal of this study was to learn more about the current scenario of culture compositions, disease prevalence, application of aqua drugs, and their effects on fish health and the environment. There are six types of cultured farms that were investigated in this experiment. The experimental area had the highest percentages of mixed culture of carp, monosex tilapia, and pangas. Some previous research also revealed similar culture compositions in the different areas of Bangladesh (Kawsar et al., 2019;Rahman et al., 2017). Current investigation showed that farmers applied the nine distinct commercial aqua drugs for various aquaculture operations, especially in health management of fishes. Besides, in pond preparation, disinfecting the culture environment, stimulating growth, and enhancing immunity number of drugs and chemicals were found in use. The primary sources of these chemicals are local animal feed and veterinary medicines stores, which is in the perimeter of the farmers and very easy to purchase. This study identified zeolite and lime as the most used chemical for pond water quality maintenance. In addition lime and zeolites are reportedly a most used chemicals in the area which is consistent to this survey (Chowdhury et al., 2012;Faruk et al., 2008;Kawsar et al., 2019;Shamsuzzaman et al., 2012).
One of the major limitations emerged in aquaculture intensification nowadays is fish diseases, which become a significant constraint in effective and sustainable aquaculture business profitability, in consequences. EUS, fin and tail rot, dropsy, bacillary necrosis, white spot, saprolegniasis, lernaeasis, and argulosis were the most common diseases and symptoms (Table 10) reported by farmers in this study. Several authors have also documented comparable circumstances in Bangladeshi aquaculture industry (Faruk et al., 2004;Amin, 2000;Mazid, 2001). According to the supplier's information, disinfectants and antibiotics were used by farmers to maintain their ponds free of pathogens and to cure several infectious diseases such as Epizootic Ulcerative Syndrome (EUS), Red spot disease, streptococcosis, ichthyophthiriasis, etc. In some cases, the farmer got about 95% recoveries from the disease condition within a short period (Rahman et al., 2017). In the current investigation, 23 disinfectant brands and 30 antibiotic brands with various trade names were found on the market. Similarly, Shamsuzzaman and Biswas (2012) identified 12 disinfectant brands and 14 antibiotic brands on the southwest coast of Bangladesh. The most often used disinfectants in the study area were commercial disinfectant such as BKC and salt potassium permanganate and bleaching powder. Rahman et al. (2017) stated that 22%, Kawsar et al. (2019) claimed that 40% of farmers of the study area used potassium permanganate and 38% were used commercial disinfectant. Antibiotics were found to be administered indiscriminately in the current investigation, although the specific causes of the disease were unknown. Some farmers did not use the prescribed treatment doses. A total of 15 antibiotics were identified and farmers were reported with irresponsible and frequent use of such drugs without approval and without knowing their effects on fish health (Faruk et al., 2021). Several issues regarding improper use of aquatic medications, such as lack of information about chemical use, sufficient dosage, form of application, and indiscriminate use of antibiotics have been reported by Hasan et al. (2020).
Several aqua drugs have been found to be used as oxygen precursors, ammonia reducers, growth promoters, antiparasitics, enzymes, and probiotics to aid digestion and keep the aquatic environment healthy. Antibiotics with six categories of other compounds, including nutritional supplements, disinfectants, saline, ammonia removal, probiotics, and pesticides, were administered by the fish farmers of Mymensingh (Faruk et al., 2021). Previously several authors found very similar outcomes in their research on aqua medicines in Bangladesh (Kawsar et al., 2019;Rahman et., al., 2017;Shamsuzzaman and Biswas, 2012;Chowdhury et al., 2012;Rahman et al., 2015).

Antibiotic exposure pathway in aquaculture
In aquaculture, as in other animal production sectors, similar strategies (e.g., vaccination and use of antibiotics) are used to control infectious diseases. Antimicrobial use in aquaculture differs from cattle farming due to the greater diversity of species, farming practices, and 26% 19% 17% 14% 6% 9% 5% 4%

Ciprofloxacin Chlortetracycline Enrofloxacin
Amoxicillin Others Figure 6. Antibiotics used in the study area. different application methods. The application of antimicrobial in aquaculture ponds has a consequence in the formation of drug-resistant bacteria repositories in aquatic species and even in the ecosystem (Schmidt et al., 2000;Akinbowale et al., 2006;Hamom et al., 2020). Antimicrobials, in whatever form these are applied in food production, these will have significant complications on human health, surrounding environment, and aquatic ecosystems (Rasul and Majumdar, 2017;Brunton et al., 2019). According to the pathways analysis (Figure 12), the two most prevalent techniques of administering antimicrobial drugs in aquaculture species are medicated feed and applying antibiotics directly to the water (immersion therapy), both of which require flock treatment of the animals (Heuer et al., 2009). Antibiotics are most often administered to aquatic animals by combining them with specially formulated feed. However, antibiotics are not efficiently metabolized by fish and are mostly execrated in the environment via faces. Seventy-five percent of antibiotics supplied to fish are estimated to be released into the aquatic environment (Burridge et al., 2010). These practices can lead to excessive usage of antimicrobials and strong selective pressure in the aquatic animals and the exposed surroundings.
According to the findings of some previous studies, effluent from antibiotic-treated cattle farms (fecal waste) may end up in aquaculture ponds since cow dung is a popular source of raw ingredients in land-based aquaculture (Jha et al., 2004), potentially introducing antibiotics into the aquaculture setting (Sobur et al., 2019;Kabir et al., 2018). Another source of antibiotics in fish farms in the study area is the dumping of poultry litter and other slaughterhouse wastes in the pond ecosystem to increase primary production. Resulting antibiotics may gain access to the pond environment, and there has recently been an increase in the incidence of food-borne illnesses caused by antibiotic-resistant bacteria (Teuber, 1999). The use of poultry litter as an aquaculture supplement may transmit certain food-borne or zoonotic bacteria and multidrug-resistant bacteria to the aquaculture setup and consequently to the consumer (Aly et al., 2009).  Combine Yucca and Bacillus sp.
Others Figure 9. Gas reducers used in the study area. Md.A. Kawsar et al. Heliyon 8 (2022) e09060 4.2. Antimicrobial resistance (AMR) in aquaculture We found a total of 30 brands of antibiotics were reported from the study area. A total of 58 antibiotics have been recorded from various parts of Bangladesh previously (Asif et al., 2021). Lulijwa et al. (2020) studied antibiotic usage in key aquaculture-producing countries, including Bangladesh, and found that 19 antibiotics are used in Bangladeshi aquaculture. This study showed that antibiotics were applied indiscreetly by fish farmers without identifying the exact causes of fish diseases. Rahman et al. (2017) described similar findings. Antibiotics used inappropriately or irrationally can lead to the development of antibiotic-resistant bacteria (Inglis V. 2000). The longer an antibiotic is   Figure 10. Enzymes used in the study area. exposed to the environment, the higher the chance of resistance developing, and administering these medications into aquatic environments allows them to survive for lengthy periods. Antibiotic residues have severe public health consequences (Table 11), including antibacterial drug resistance, hypersensitivity reactions, mutagenicity, carcinogenicity, bone marrow suppression, teratogenicity, and disruption of normal gut flora (Schar et al., 2020;Okocha et al., 2018;Miranda et al., 2018).

Options for risk management
The most efficient way to manage and control the emergence and expansion of antimicrobial resistance is to minimize the demand for antibacterial treatment (Moges et al., 2014). A federal administrative framework is necessary for antibacterial agent licensing, approval, monitoring, and regulation in all countries where antimicrobial agents are applied in aquatic biota. Production planning should include stocking strategies and management measures to prevent the invasion of germs and the spread of infectious diseases. As Bangladesh is a leading aquaculture producer, the government should place a high priority on the control and monitoring of aquaculture drugs and chemicals at the field level. When it comes to the usage of aqua drugs and antibiotics, there are a few guidelines to follow, including (i) constantly endeavor to enhance the pond ecosystem, (ii) aqua drugs and antibiotics should only be used when it is essential, (iii) use antimicrobials only for bacterial infection,  Figure 12. The possible exposure pathway of antibiotics through a hypothetically illustrated aquaculture setup. Different sources like raw materials of the feed, some intentional approaches of the farmers, and various waste disposals input the antibiotics into the setup, which afterward affect the local environment and public health through contaminated water sources and sediments, including consumption and handling of the aquaculture end products and in consequences, antimicrobial resistance becomes developed into different ecological compartments (Santos and Ramos, 2018;Ngogang et al., 2021;Ullah et al., 2020;Thai et al., 2018;Han et al., 2017;Chowdhury et al., 2015;Pruden et al., 2013).  Figure 11. Antiparasitics used in the study area. Md.A. Kawsar et al. Heliyon 8 (2022) e09060 (iv) use an antimicrobial to which the pathogens are susceptible, (v) arrange training on Good Aquaculture Practices (GAP) for farmers, (vi) Aqua chemicals should be handled with caution since they can be harmful to public health, (vii) use appropriate dose, (viii) Minimum use of chemical is the best alternative to reduce adverse effect.

Conclusion
Considering the tremendous expansion and significance of the aquaculture sector in several areas of Bangladesh, the study sought to ascertain the current state of aquaculture inputs like chemicals and antimicrobial substance use in aquatic animal care and identified several challenges when it comes to using chemicals, including a lack of understanding of the compounds, their unregulated uses, and application methods. Significant measures are required to combat the establishment and spread of indiscriminate chemical usage and antimicrobial resistance in aquaculture. The efficacy of various aqua products in field trials and fish disease diagnosis based on signs, symptoms, and eye assessment may be considered a significant research gap in this study. However, to reduce the detrimental effects of drugs used in aquafarming, government policymakers, fisheries experts, researchers, farmers, entrepreneurs, the pharmaceutical industry, and scientists should collaborate to address the challenges.

Author contribution statement
Md. Abu Kawsar: Conceived and designed the experiments; Performed the experiments; Analyzed and interpreted the data; Contributed reagents, materials, analysis tools or data; Wrote the paper.
Mamun Mia: Performed the experiments. Tofael Ahmed Sumon: Analyzed and interpreted the data; Wrote the paper.

Funding statement
This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.

Data availability statement
Data included in article/supplementary material/referenced in article.

Declaration of interests statement
The authors declare no conflict of interest.

Additional information
No additional information is available for this paper.